Phloroglucinol (1,3,5-trihydroxybenzene) and its derivatives are widely used in commerce. Phloroglucinol and its derivatives, e.g., trimethylphloroglucinol, are used as pharmaceutical agents, e.g., as antispasmodics. Phloroglucinol is used as a starting material or intermediate in pharmaceutical, microbicide, and other organic syntheses. Phloroglucinol is used as a stain for microscopy samples that contain lignin (e.g., wood samples), and it is used in the manufacture of dyes, including leather, textile, and hair dyes. It is used in the manufacture of adhesives and as an epoxy resin curing agent, and in the preparation of explosives, e.g., the thermally- and shock-stable high explosive, 1,3,4-triamino-2,4,6-trinitrobenzene (TATB). Phloroglucinol also functions as an antioxidant, stabilizer, and corrosion resistance agent, and is utilized as a coupling agent for photosensitive duplicating paper, as a substitute for silver iodide in rain-making, as a bone sample decalcifying agent, and as a floral preservative. Phloroglucinol can also be converted to resorcinol by catalytic hydrogenation.
Resorcinol (1,3-dihydroxybenzene) is a particularly useful derivative of phloroglucinol, although resorcinol is not currently produced by that route. As is phloroglucinol, resorcinol is used in the manufacture of dyes and adhesives, and as an epoxy resin curing agent; and it is used as a starting material or intermediate in pharmaceutical and other organic syntheses. Resorcinol and its derivatives are further commonly used, either alone or with other active ingredients such as sulfur, in cosmetics and in topical skin medicaments for treatment of conditions including acne, dandruff, eczema, and psoriasis, functioning, in part, as an antiseptic and antipruritic. Resorcinol is also used as a cross-linking agent for neoprene, as a tack-enhancing agent in rubber compositions, in bonding agents for organic polymers (e.g., melamine and rubber), and in the fabrication of fibrous and other composite materials. Resorcinol is used: in the manufacture of resins and resin adhesives, e.g., both as a monomer and as a UV absorbing agent; in the manufacture of explosives, e.g., energetic compounds such as styphnic acid (2,4,6-trinitrobenzene-1,3-diol) and heavy metal styphnates; and in the synthesis of diazo dyes, plasticizers, hexyl resorcinol, and p-aminosalicylic acid.
The most common of the resorcinol-based resins are resorcinol-aldehyde and resorcinol-phenol-aldehyde resins. These types of resorcinol-based resins are used, for example, as resin adhesives, composite material matrices, and as starting materials for rayon and nylon production. Examples of composite materials include resorcinol-formaldehyde carbon (or other organic) particle hydrogels, aerogels, and xerogels, which are useful, e.g., as matrix materials for metallic and organometallic catalysts. Resorcinol-formaldehyde resins and particulate composites therewith are also used in dentistry as a root canal filling material.
Resorcinol-aldehyde resin adhesives are especially useful in applications requiring high bond strength, including, e.g.: wooden trusses, joists, barrels, and boats; and aircraft. Modified resorcinol-aldehyde resin adhesives are also used as biological wound sealant compositions both on topical wounds and on internal wounds or surgical cuts, e.g., vascular incisions. This is often done in military field medicine, e.g., to minimize environmental exposure, reduce bleeding and fluid loss, and speed the healing process. Such modified resin adhesives include, e.g., gelatin-resorcinol-formaldehyde and gelatin-resorcinol-glutaraldehyde compositions, wherein the aldehyde may be maintained separately from, and later mixed with, the resorcinol-gelatin composition to form the sealant when needed.
Currently, both phloroglucinol and resorcinol are commercially produced by chemical organic synthesis using caustics and high temperatures, beginning with petroleum-derived starting materials and creating much environmentally problematic waste.
As a result, it would be an improvement in the art to provide more efficient and cleaner processes for the production of these valuable compounds. One possible solution might be to provide a biosynthetic route for production of phloroglucinol, with an optional hydrogenation of the biosynthetic phloroglucinol to resorcinol. Biosynthetic production of compounds related to phloroglucinol has been reported in plants, algae, and microbes, e.g.: acetyl phloroglucinols from Pseudomonas spp.; hyperforins, hyperfoliatins, hyperjovinols, and hyperatomarins from Hypericum spp.; pallidusol, dehydropallidusol, pallidol, mallopallidol, and homomallopallidol from Mallotus spp.; garcinielliptones from Garcinia spp.; flavaspidic acids from Dryopteris spp.; macrocarpals and sideroxylonals from Eucalyptus spp.; 1,3,5-trimethoxybenzene from Rosa spp.; as well as phloroglucinol-containing glycosides and phlorotannins.
However, production of phloroglucinol is reported in such plants and microbes as merely a degradation product of more complex, and thus less abundant and/or more costly, starting materials. See, e.g.: L. Schoefer et al., Appl. Environ. Microbiol. 70(10):6131-37 (2004); D. Baas & J. Rétey, Eur. J. Biochem. 265:896-901 (1999). In addition, microbial biosynthetic production of di-acetyl phloroglucinols has been proposed as a means for improving the anti-fungal activity of recombinant bacteria to be released into the agricultural environment as biocontrol agents against phytopathogens. See U.S. Pat. No. 6,051,383, Thomashow et al., issued Apr. 18, 2000; and M. G. Bangera & L. S. Thomashow, J Bact. 181(10):3155-63 (1999). Yet, a route of anabolic biosynthetic production of phloroglucinol, e.g., from inexpensive starting materials such as glucose, is not shown.
Recently, an alternate route (see FIG. 2) to phloroglucinol (1a) has been elaborated, which involves microbe-catalyzed synthesis of triacetic acid lactone (3a) from glucose; however, it has been found that multiple chemical steps are needed to convert triacetic acid lactone (3a) into phloroglucinol (1a). See W. Zha et al., J. Am. Chem. Soc. 126(14):4534-35 (2004); and C. A. Hansen & J. W. Frost, J. Am. Chem. Soc. 124(21):5926-27 (2002). Thus, this route is at best a partly biosynthetic, partly chemosynthetic pathway.
Thus, to date, no fully biosynthetic route useful for commercial production of phloroglucinol per se (1,3,5-trihydroxybenzene) has been reported. No enzymes or encoding genes that catalyze the formation of phloroglucinol per se have been identified.